O-aryl, O-alkyl, O-alkenyl and O-alkynylrapamycin derivatives

ABSTRACT

O-Aryl, O-alkyl, O-alkenyl and O-alkynylrapamycin derivatives of the general structural Formula I: ##STR1## have been prepared from suitable precursors by alkylation and/or arylation at C-42 and/or C-31. These compounds are useful in a mammalian host for the treatment of autoimmune diseases and diseases of inflammation, infectious diseases, the prevention of rejection of foreign organ transplants and the treatment of solid tumors.

SUMMARY OF THE INVENTION

The present invention is related to O-aryl, O-alkyl, O-alkenyl andO-alkynylrapamycin derivatives which are useful in a mammalian host forthe treatment of autoimmune diseases (such as juvenile-onset orrecent-onset diabetes mellitus, multiple sclerosis, rheumatoidarthritis, liver disease, posterior uveitis, allergic encephalomyelitis,and glomerulonephritis), diseases of inflammation, infectious diseases(particularly fungal infections), the prevention of rejection of foreignorgan transplants, e.g. bone marrow, kidney, liver, heart, skin,small-bowel, and pancreatic-islet-cell transplants, and the treatment ofsolid tumors.

More particularly, this invention relates to compounds of the generalstructural Formula I: ##STR2## wherein R¹ and R² are hereinafterdefined.

This invention also relates to pharmaceutical compositions containingthe compounds, and to a method of use of the present compounds and otheragents for the treatment and prevention of certain afflictions, diseasesand illnesses.

BACKGROUND OF THE INVENTION

Rapamycin characterized by Findlay and coworkers in 1978 is a35-membered macrolide isolated from S. hygroscopicus (Can. J. Chem.,1978, 56, 2491, J. Antibiotics, 1975, 28, 721, U.S. Pat. No. 3,929,992,issued Dec. 30, 1975, U.S. Pat. No. 3,993,749, issued Nov. 23, 1975.Rapamycin has been found to have antifungal activity, particularlyagainst Candida albicans, both in vitro and in vivo (J. Antibiotics,1978, 31, 539).

Rapamycin alone (U.S. Pat. No. 4,885,171) or in combination withpicibanil (U.S. Pat. No. 4,401,653) has been shown to have antitumoractivity. R. Martel et al. (Can, J. Physiol. Pharmacol, 55, 48 (1977)disclosed that rapamycin is effective in an experimental allergicencephalomyelitis model, a model for multiple sclerosis; in an adjuvantarthritis model, a model for rheumatoid arthritis; and effectivelyinhibited the formation of IgE-like antibodies.

The immunosuppressive effects of rapamycin have been disclosed (FASEB 3,3411 (1989); Med. Sci. Res., 1989, 17, 877). Cyclosporin A and FK-506,other macrocyclic molecules, also have been shown to be effective asimmunosuppressive agents, therefore useful in preventing transplantrejection (FASEB 3, 3411 (1989); FASEB 3, 5256 (1989); and Lancet 1183(1978)).

Fujisawa United States patents (U.S. Pat. No. 4,929,611, issued May 29,1990 and U.S. Pat. No. 4,956,352, issued Sept. 11, 1990) disclose theuse of FK-506-type compounds in treating resistance to transplantation.A Sandoz European patent application (EPO Publication No. 0,315,978)discloses the use of FR-900506 and related compounds in the topicaltreatment of inflammatory and hyperproliferative skin diseases and ofcutaneous manifestations of immunologically-mediated illness. A FisonsWorld patent application (PCT Publication WO 90/14826) discloses the useof FR-900506 and related compounds in the treatment of reversibleobstructive airways disease, particularly asthma. A Fujisawa Europeanpatent application (EPO Publication No. 0,423,714) discloses the use ofFK-506 and derivatives as hair revitalizing agents. Various studies havesuggested the efficacy of FK-506 in the treatment of a number ofailments, including rheumatoid arthitis (C. Arita, et al., Clincial exp.Immunol., 1990, 82, 456-461; N. Inamura, et al., Clin. Immunol.Immunopathol. 1988, 46, 82-90), recent-onset diabetes (N. Murase, etal., Diabetes., 1990, 39, 1584-86; N. Murase, et al., Lancet, 1990, 336,373-74), posterior uveitis (H. Kawashima, Invest. Ophthalmul, Vis. Sci.,1988, 29, 1265-71), hepatic injury associated with ischemia (M. Sake, etal., Life Sci., 1990, 47, 687-91) allergic encephalomyelitis (K,Deguchi, et al., Brain Nerve, 1990, 42, 391-97), glomerulonephritis (J.McCauley, et al., Lancet, 1990, 335, 674), systemic lupus erythematosus(K. Takabayashi, et al., Clin. Immunol, Immunopathol., 1989, 51,110-117), multidrug resistance (M. Naito, et al., Cancer Chemother,Pharmacol., 1992, 29, 195-200), inflammation of mucosa and blood vessels(PCT Publication WO 91/17754), cytomegalovirus infection (UK PublicationGB 2,247,620A), and idiopathic thrombocytophenic purpura and Basedow'sdisease (PCT Publication WO 91/19495).

Mono- and diacylated derivatives of rapamycin (esterified at the 31 and42 positions) have been shown to be useful as antifungal agents (U.S.Pat. No. 4,316,885) and used to make water soluble prodrugs of rapamycin(U.S. Pat. No. 4,650,803). Reduction products of rapamycin have beenprepared (U.S. Pat. Nos. 5,102,876 and 5,138,051). Derivatives ofrapamycin at the 31 and 42 positions which have been disclosed include:carboxylic acid esters (PCT Patent Publication WO92/05179); carbamates(U.S. Pat. No. 5.118.678); amide esters (U.S. Pat. No. 5,118,677);fluorinated esters (U.S. Pat. No. 5,100,883); acetals (U.S. Pat. No.5,151,413); and silyl ethers (U.S. Pat. No. 5,120,842). In addition,bicyclic derivatives of rapamycin connected via the 31, 42 positions(U.S. Pat. No. 5,120,725) and rapamycin dimers connected via the 42position (U.S. No. Pat. 5,120,727) have been disclosed. Variousaryl(lower alkyl) and heteroaryl derivatives of FK-506 type compoundshave also been disclosed (UK Patent Publication No. GB 2,245,891A).O-Aryl, O-alkyl, O-alkenyl and O-alkynyl derivatives of FK-506 typecompounds will have been disclosed (EPO Patent Publication No.0,515,071).

DETAILED DESCRIPTION OF THE INVENTION A. Scope of the Invention

The novel compound of this invention has structural Formula I: ##STR3##or a pharmaceutically acceptable salt thereof, wherein: R¹ and R² areindependently selected from:

(1) hydrogen;

(2) phenyl;

(3) substituted phenyl in which the substituents are X, Y and Z;

(4) 1- or 2- naphthyl;

(5) substituted 1- or 2- naphthyl in which the substituents are X, Y andZ;

(6) biphenyl;

(7) substituted biphenyl in which the substituents are X, Y and Z;

(8) C₁₋₁₀ alkyl;

(9) substituted C₁₋₁₀ alkyl in which one or more substituent(s) is(are)selected from:

(a) hydroxy,

(b) oxo,

(c) C₁₋₆ -alkoxy,

(d) phenyl-C₁₋₃ alkoxy,

(e) substituted phenyl-C₁₋₃ alkoxy, in which the substituents on phenylare X, Y and Z,

(f) --OCO--C₁₋₆ alkyl,

(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are independently selected from

(i) hydrogen,

(ii) C₁₋₁₀ alkyl unsubstituted or substituted with one or more of thesubstituent(s) selected from:

(a') phenyl, which is unsubstituted or substituted with X, Y and Z,

(b') --OH,

(c') C₁₋₆ alkoxy,

(d') --CO₂ H,

(e') --CO₂ --C₁₋₆ alkyl,

(f') --C₃₋₇ cycloalkyl, and

(g') --OR¹¹,

(iii)C₃₋₁₀ alkenyl unsubstituted or substituted with one or more of thesubstituent(s) selected from:

(a') phenyl, which is unsubstituted or substituted with X, Y and Z,

(b') --OH,

(c') C₁₋₆ alkoxy,

(d') CO₂ H,

(e') --CO₂ --C₁₋₆ alkyl,

(f') --C₃₋₇ cycloalkyl, and

(g') --OR¹¹,

(iv) or where R⁶ and R⁷ and the N to which they are attached can form anunsubstituted or substituted 3-7-membered saturated heterocyclic ringwhich can include one or two additional heteroatoms independentlyselected from the group consisting of O S(O)_(p), NR¹⁴, wherein R¹⁴ ishydrogen or C₁₋₆ alkyl unsubstituted or substituted by phenyl, and p is0, 1 or 2, the ring being selected from the group consisting of:aziridine, morpholine, thiomorpholine, thiomorpholine-oxide,thiomorpholine-dioxide, piperidine, pyrrolidine, and piperazine,

(h) --NR⁶ CO--C₁₋₆ alkyl-R⁷, wherein R⁶ is as defined above,

(i) --NR⁶ CO₂ --C₁₋₆ alkyl-R⁷,

(j) --NR⁶ CONR⁶ R⁷,

(k) --OCONR R⁶ R⁷,

(l) --COOR⁶,

(m) --CHO,

(n) phenyl,

(o) substituted phenyl in which the substituents are X, Y and Z,

(p) phenyloxy,

(q) substituted phenyloxy in which the substituents are X, Y and Z,

(r) 1- or 2- naphthyl,

(s) substituted 1- or 2- naphthyl in which the substituents are X, Y andZ,

(t) biphenyl

(u) substituted biphenyl in which the substituents are X, Y and Z.

(v) --OR¹¹, and

(w) --S(O)_(p) --C₁₋₆ alkyl;

(10) C₃₋₁₀ alkenyl;

(11) substituted C₃₋₁₀ alkenyl in which one or more substituent(s)is(are) selected from:

(a) hydroxy,

(b) oxo,

(c) C₁₋₆ alkoxy,

(d) phenyl-C₁₋₃ alkoxy,

(e) substituted phenyl-C₁₋₃ alkoxy, in which the substituents on phenylare X, Y and Z,

(f) --OCO--C₁₋₆ alkyl,

(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above

(h) --NR⁶ CO--C₁₋₆ alkyl, wherein R⁶ is as defined above,

(i) --COOR⁶, wherein R⁶ is as defined above,

(j) --CHO,

(k) phenyl,

(l) substituted phenyl in which the substituents are X, Y and Z,

(m) 1- or 2-naphthyl,

(n) substituted 1- or 2-naphthyl in which the substituents are X, Y andZ,

(o) biphenyl,

(p) substituted biphenyl in which the substituents are X, Y and Z,

(q) --OR¹¹, and

(r) --S(O)_(p) --C₁₋₆ alkyl;

(12) C₃₋₁₀ alkyl;

(13) substituted C₃₋₁₀ alkynyl in which one or more substituent(s)is(are) selected from:

(a) hydroxy,

(b) oxo,

(c) C₁₋₆ alkoxy,

(d) phenyl-C₁₋₃ alkoxy,

(e) substituted phenyl-C₁₋₃ alkoxy, in which the substituents on phenylare X, Y and Z,

(f) --OCO--C₁₋₆ alkyl,

(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above,

(h) --NR⁶ CO--C₁₋₆ alkyl, wherein R⁶ is as defined above,

(i) --COOR⁶ is as defined above,

(j) --CHO,

(k) phenyl,

(1) substituted phenyl in which the substituents are X, Y and Z,

(m) 1- or 2-naphthyl,

(n) substituted 1- or 2-naphthyl in which the substituents are X, Y andZ,

(o) biphenyl,

(p) substituted biphenyl in which the substituents are X, Y and Z, and

(q) --OR¹¹ ;

with the proviso that R¹ and R² are not simultaneously hydrogen;

R¹¹ is selected from:

(a) --PO(OH)O⁻ M⁺, wherein M⁺ is a positively charged inorganic ororganic counterion,

(b) --SO₃ ⁻ M⁺,

(c) --CO(CH₂)_(q) CO₂ ⁻ M⁺, wherein q is 1-3, and

(d) --CO--C₁₋₆ alkyl--NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above andthe alkyl is unsubstituted or substituted with one or more substituentsselected from:

(i) hydroxy,

(ii) C₁₋₆ alkoxy,

(iii) --NR¹⁶ R¹⁷, wherein R¹⁶ and R¹⁷ are independently selected from:

(a') hydrogen, and

(b') C₁₋₆ alkyl,

(iv) --COOR⁶, wherein R⁶ is as defined above,

(v) phenyl,

(vi) substituted phenyl in which the substituents are X, Y and Z,

(vii) --SH, and

(viii) --S--C₁₋₆ alkyl;

X, Y and Z independently are selected from:

(a) hydrogen,

(b) C₁₋₇ alkyl,

(c) C₂₋₆ alkenyl,

(d) halogen,

(e) --(CH₂)_(m) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above, and mis 0 to 2,

(f) --CN,

(g) --CHO,

(h) --CF₃,

(i) --SR⁸, wherein R⁸ is hydrogen, C₁₋₆ alkyl, trifluoromethyl, orphenyl,

(j) --SOR⁸, wherein R⁸ is as defined above,

(k) --SO₂ R⁸, wherein R⁸ is as defined above,

(l) --CONR⁶ R⁷, wherein R⁶ and R⁷ are as defined above,

(m) R⁹ O(CH₂)_(m) - wherein R⁹ is hydrogen, C₁₋₃ alkyl, hydroxy-C₂₋₃alkyl, trifluoromethyl, phenyl or naphthyl and m is as defined above,

(n) --CH(OR¹²)(OR¹³), wherein R¹² and R¹³ are C₁₋₃ alkyl or takentogether form an ethyl or propyl bridge,

(o) ##STR4## wherein R⁹ and m are as defined above, and (p) ##STR5##wherein R⁹ and m are as defined above, and (q) --OR¹¹ ;

or any two of adjacent X, Y and Z can be joined to form a ring having 5,6 or 7 ring atoms, said ring atoms comprising 1 or 2 oxygen atoms, theremaining ring atoms being carbon, selected from the group consistingof: dioxolanyl, dihydrofuranyl, dihydropyranyl, and dioxanyl.

The compounds of the present invention have asymmetric centers and thisinvention includes all of the optical isomers and mixtures thereof.

In addition compounds with carbon-carbon double bonds may occur in Z-and E- forms with all isomeric forms of the compounds being included inthe present invention.

When any variable (e.g., alkyl, aryl, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹, etc.)occurs more than one time in any variable or in Formula I, itsdefinition on each occurrence is independent of its definition at everyother occurrence.

As used herein, the term "alkyll" includes those alkyl groups of adesignated number of carbon atoms of either a straight, branched, orcyclic configuration. Examples of "alkyl" include methyl, ethyl, propyl,isopropyl, butyl, sec-and tert-butyl, pentyl, hexyl, heptyl,cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl,norbornyl, and the like. "Alkoxy" represents an alkyl group of indicatednumber of carbon atoms attached through an oxygen bridge, such asmethoxy, ethoxy, propoxy, butoxy and pentoxy.

"Alkanoyl" is intended to include those alkylcarbonyl groups ofspecified number of carbon atoms, which are exemplified by formyl,acetyl, propanoyl and butyryl; "alkanoyloxy" is intended to includethose alkylcarbonyl groups of specified number of carbon atoms attachedthrough an oxygen bridge, which are exemplified by formyloxy, acetoxy,propionoyloxy, and butyryloxy. "Alkenyl" is intended to includehydrocarbon chains of a specified number of carbon atoms of either astraight- or branched-configuration and at least one unsaturation, whichmay occur at any point along the chain, such as ethenyl, propenyl,butenyl, pentenyl, dimethyl pentenyl, and the like, and includes E and Zforms, where applicable; and "arylalkyl" represents aryl groups asherein defined which are attached through a straight or branched chainalkyl group of from one to six carbon atoms, such as, for example,benzyl, phenethyl, 3,3-diphenylpropyl, and the like. "Halogen", as usedherein, means fluoro, chloro, bromo and iodo.

As will be understood by those skilled in the art, pharmaceuticallyacceptable salts include, but are not limited to salts with inorganicacids such as hydrochloride, sulfate, phosphate, diphosphate,hydrobromide, and nitrate or salts with an organic acid such as malate,maleate, fumarate, tartrate, succinate, citrate, acetate, lactate,methanesulfonate, p-toluenesulfonate or palmoate, salicylate andstearate. Similarly pharmaceutically acceptable cations include, but arenot limited to sodium, potassium, calcium, aluminum, lithium andammonium (especially ammonium salts with -amines of the formula HNR⁶R⁷). One embodiment of the present invention encompasses the compoundsof Formula I wherein:

R¹ and R² are independently selected from:

(1) hydrogen;

(2) methyl;

(3) phenyl;

(4) substituted phenyl in which the substituents are X, Y and Z;

(5) 1- or 2- naphthyl;

(6) substituted 1- or 2- naphthyl in which the substituents are X, Y andZ;

(7) biphenyl; and

(8) substituted and biphenyl in which the substituents are X, Y and Z;

with the proviso that R¹ and R² are not simultaneously hydrogen;

X, Y and Z are independently, selected from:

(a) hydrogen,

(b) C₁₋₇ alkyl,

(c) C₂₋₆ alkenyl,

(d) halogen,

(e) --(CH₂)_(m) --NR⁶ R⁷, wherein R6 and R are, independently selectedfrom

(i) hydrogen, or

(ii) C₁₋₆ alkyl unsubstituted or substituted with phenyl, and m is 0 to2,

(f) --CN,

(g) --CHO,

(h) --CF₃,

(i) --SR⁸, wherein R⁸ is hydrogen, C₁₋₆ alkyl, trifluoromethyl, orphenyl,

(j) --SOR⁸, wherein R⁸ is as defined above,

(k) --SO₂ R⁸, wherein R⁸ is as defined above,

(l) --CONR⁶ R⁷, wherein R⁶ and R⁷ are as defined above,

(m) R⁹ O(CH₂)_(m) - wherein R⁹ is hydrogen, C₁₋₃ alkyl, hydroxy-C₂₋₃alkyl, trifluoromethyl, phenyl or naphthyl and m is as defined above,

(n) --CH(OR¹²)(OR¹³), wherein R¹² and R¹³ are C₁₋₃ alkyl or takentogether form an ethyl or propyl bridge,

(o) ##STR6## wherein R⁹ and m are as defined above, and (p) ##STR7##wherein R⁹ and m are as defined above, and (q) --OR¹¹ ;

or any two of adjacent X, Y and Z can be joined to form a ring having5,6 or 7 ring atoms, said ring atoms comprising 1 or 2 oxygen atoms, theremaining ring atoms being carbon, selected from the group consistingof: dioxolanyl, dihydrofuranyl, dihydropyranyl, and dioxanyl.

Another embodiment of the present invention encompasses the compounds ofFormula I wherein:

R¹ and R² are independently selected from:

(1) hydrogen;

(2) C₁₋₁₀ alkyl; (3) substituted C₁₋₁₀ alkyl in which one or moresubstituent(s) is(are) selected from:

(a) hydroxy,

(b) oxo,

(c) C₁₋₆ -alkoxy,

(d) phenyl-C₁₋₃ alkoxy,

(e) substituted phenyl-C₁₋₃ alkoxy, in which the substituents on phenylare X, Y and Z,

(f) --OCO--C₁₋₆ alkyl,

(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are independently selected from

(i) hydrogen,

(ii) C₁₋₁₀ alkyl unsubstituted or substituted with one or more of thesubstituent(s) selected from:

(a') phenyl, which is unsubstituted or substituted with X, Y and Z,

(b') --OH,

(c') C₁₋₆ alkoxy,

(d') --CO₂ H,

(e') --CO₂ --C₁₋₆ alkyl,

(f') --C₃₁₄ 7 cycloalkyl, and

(g') --OR¹¹,

(iii)C₃₋₁₀ alkenyl unsubstituted or substituted with one or more of thesubstituent(s) selected from:

(a') phenyl, which is unsubstituted or substituted with X, Y and Z,

(b') --OH,

(c') C₁₋₆ alkoxy,

(d') --CO₂ H,

(e') --CO₂ --C₁₋₆ alkyl,

(f') --C₃₋₇ cycloalkyl, and

(g') --OR¹¹,

(iv)or where R⁶ and R⁷ and the N to which they are attached can form anunsubstituted or substituted 3-7-membered saturated heterocyclic ringwhich can include one or two additional heteroatoms independentlyselected from the group consisting of O S(O)_(p), NR¹⁴, wherein R¹⁴ ishydrogen or C₁₋₆ alkyl unsubstituted or substituted by phenyl, and p is0, 1 or 2, the ring being selected from the group consisting of:aziridine, morpholine, thiomorpholine, thiomorpholine-oxide,thiomorpholine-dioxide, piperidine, pyrrolidine, and piperazine,

(h) --NR⁶ CO--C₁₋₆ alkyl-R⁷, wherein R⁶ is as defined above,

(i) --NR⁶ CO₂ --C₁₋₆ alkyl-R⁷,

(j) --NR⁶ CONR⁶ R⁷,

(k) --OCONR⁶ R⁷,

(l) --COOR⁶,

(m) --CHO,

(n) phenyl,

(o) substituted phenyl in which the substituents are X, Y and Z,

(p) phenyloxy,

(q) substituted phenyloxy in which the substituents are X, Y and Z,

(r) 1- or 2- naphthyl,

(s) substituted 1- or 2- naphthyl in which the substituents are X, Y andZ,

(t) biphenyl

(u) substituted biphenyl in which the substituents are X, Y and Z;

(v) --OR¹¹, and

(w) --S(O)_(p) --C₁₋₆ alkyl;

(4) C₃₋₁₀ alkenyl;

(5) substituted C₃₋₁₀ alkenyl in which one or more substituent(s)is(are) selected from:

(a) hydroxy,

(b) oxo,

(c) C₁₋₆ alkoxy,

(d) phenyl-C₁₋₃ alkoxy,

(e) substituted phenyl-C₁₋₃ alkoxy, in which the substituents on phenylare X, Y and Z,

(f) --OCO--C₁₋₆ alkyl,

(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above

(h) --NR⁶ CO--C₁₋₆ alkyl, wherein R⁶ is as defined above,

(i) --COOR⁶, wherein R⁶ is as defined above,

(j) --CHO,

(k) phenyl,

(l) substituted phenyl in which the substituents are X, Y and Z,

(m) 1- or 2-naphthyl,

(n) substituted 1- or 2-naphthyl in which the substituents are X, Y andZ,

(o) biphenyl,

(p) substituted biphenyl in which the substituents are X, Y and Z,

(q) --OR¹¹, and

(r) --S(O)_(p) --C₁₋₆ alkyl;

(6) C₃₋₁₀ alkynyl;

(7) substituted C₃₋₁₀ alkynyl in which one or more substituent(s)is(are) selected from:

(a) hydroxy,

(b) oxo,

(c) C₁₋₆ alkoxy,

(d) phenyl-C₁₋₃ alkoxy,

(e) substituted phenyl-C₁₋₃ alkoxy, in which the substituents on phenylare X, Y and Z,

(f) --OCO-C₁₋₆ alkyl,

(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above,

(h) --NR⁶ CO-C₁₋₆ alkyl, wherein R⁶ is as defined above,

(i) --COOR⁶, wherein R⁶ is as defined above,

(j) --CHO,

(k) phenyl,

(l) substituted phenyl in which the substituents are X, Y and Z,

(m) 1- or 2-naphthyl,

(n) substituted 1- or 2-naphthyl in which the substituents are X, Y andZ,

(o) biphenyl,

(p) substituted biphenyl in which the substituents are X, Y and Z, and

(q) --OR¹¹ ;

with the proviso that R¹ and R² are not simultaneously hydrogen;

R¹¹ is selected from:

(a) --PO(OH)O⁻ M⁺, wherein M⁺ is a positively charged inorganic ororganic counterion,

(b) --SO₃ ⁻ M⁺,

(c) --CO(CH₂)_(q) CO₂ ⁻ M⁺, wherein q is 1-3, and

(d) --CO-C₁₋₆ alkyl-NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above andthe alkyl is unsubstituted or substituted with one or more substituentsselected from:

(i) hydroxy,

(ii) C₁₋₆ alkoxy,

(iii) --NR¹⁶ R¹⁷, wherein R¹⁶ and R¹⁷ are independently selected from:

(a') hydrogen, and

(b') C₁₋₆ alkyl,

(iv) --COOR⁶, wherein R⁶ is as defined above,

(v) phenyl,

(vi) substituted phenyl in which the substituents are X, Y and Z,

(vii) --SH, and

(viii) --S-C₁₋₆ alkyl;

X, Y and Z independently are selected from:

(a) hydrogen,

(b) C₁₋₇ alkyl,

(c) C₂₋₆ alkenyl,

(d) halogen,

(e) --(CH₂)_(m) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above, and mis 0 to 2,

(f) --CN,

(g) --CHO,

(h) --CF₃,

(i) --SR⁸, wherein R⁸ is hydrogen, C₁₋₆ alkyl, trifluoromethyl, orphenyl,

(j) --SOR⁸, wherein R⁸ is as defined above,

(k) --SO₂ R⁸, wherein R⁸ is as defined above,

(l) --CONR⁶ R⁷, wherein R⁶ and R⁷ are as defined above,

(m) R⁹ O(CH₂)_(m) - wherein R⁹ is hydrogen, C₁₋₃ alkyl, hydroxy-C₂₋₃alkyl, trifluoromethyl, phenyl or naphthyl and m is as defined above,

(n) --CH(OR¹²)(OR¹³), wherein R¹² and R¹³ are C₁₋₃ alkyl or takentogether form an ethyl or propyl bridge,

(o) ##STR8## wherein R⁹ and m are as defined above, and (p) ##STR9##wherein R⁹ and m are as defined above, and (q) --OR¹¹ ;

or any two of adjacent X, Y and Z can be joined to form a ring having 5,6 or 7 ring atoms, said ring atoms comprising 1 or 2 oxygen atoms, theremaining ring atoms being carbon, selected from the group consistingof: dioxolanyl, dihydrofuranyl, dihydropyranyl, and dioxanyl.

B. Preparation of Compounds Within the Scope of the Present Invention

The starting material for the preparation of the compounds of thisinvention is rapamycin: ##STR10##

The production and characterization of rapamycin is well know in theliterature (see U.S. Pat. No. 3,929,992 issued Dec. 30, 1975; U.S. Pat.No. 3,993,749 issued Nov. 23, 1976). Analogs of rapamycin, such as30-desmethylrapamycin (see PCT Patent Publication WO 92/14737) may alsobe employed as starting material to give analagous derivatives. Thenovel processes for preparing the novel compounds of the presentinvention are illustrated as follows, wherein R¹ and R² are as definedabove unless otherwise indicated. It will be readily apparent to one ofordinary skill in the art reviewing the synthetic route depicted belowthat other compounds within Formula I can be synthesized by substitutionof appropriate reactants and agents in the synthesis shown below.##STR11##

Reaction Scheme A

Protection of the C-31 and/or the C-42 hydroxy group may be accomplishedby methods known in the prior art for rapamycin (see e.g. U.S. Pat. No.5,120,842) such as by treatment with: 2,6-lutidine and triisopropylsilyltrifluoromethanesulfonate in a solution of methylene chloride;2,6-lutidine and t-butyldimethylsilyl trifluoromethanesulfonate in asolution of methylene chloride; pyridine and acetic anhydride in asolution of methylene chloride; pyridine and benzoyl chloride in asolution of methylene chloride; pyridine and p-nitrobenzoyl chloride ina solution of methylene chloride; imidazole and t-butyldiphenylsilylchloride in a solution of methylene chloride; and the like. For example,as shown in Reaction Scheme A, rapamycin may be protected at C-42 as thet-butyldimethylsilyl ether by treatment with one equivalent oft-butyldimethylsilyl trifluoromethanesulfonate in methylene chloride togive the C-42-di-O-TBDMS macrolide. Treatment with two equivalents ofTBDMS triflate followed by treatment with acetic acid ortoluene-sulfonic acid in methanol results in selective removal of theC-42 ether to give the C-31-O-TBDMS macrolide.

Reaction Scheme B

As shown in Reaction Scheme B, a solution of rapamycin in an inertorganic solvent such as methylene chloride, benzene, toluene,chloroform, or the like or mixtures thereof may be treated with atriarylbismuth diacetate reagent (wherein R¹ is aryl) (preparedimmediately prior to use by the addition of acetic acid to a suspensionof a triarylbismuth carbonate in an inert organic solvent such asmethylene chloride, chloroform or the like or mixtures thereof) in thepresence of a catalytic amount of copper(II) acetate at a temperature of20°-50° C., preferably room temperature, for a period of one hour toseven days, preferably one day, to give the 42-O-aryl rapamycin and/orthe 31, 42-di-O-aryl rapamycin. Alternatively, the triarylbismuth(V)reagent may be prepared by treatment of a triarylbismuthine with asuitable oxidant such as peracetic acid, iodobenzene diacetate,bis(trifluoroacetoxy)iodobenzene and the like in an inert solvent suchas methylene chloride, chloroform, benzene, toluene and the like. Thetriarylbismuth(V) reagent may be used without purification or may bepurified by silica gel chromatography. Triarylbismuthines may beprepared by the reaction of an appropriate aryl Grignard reagent withbismuth trichloride in an inert organic solvent such as tetrahydrofuran,diethyl ether, or 1,4-dioxane, or mixtures thereof, at or near roomtemperature for a period of 1 to 48 hours. General procedures for thepreparation and use of triaryl bismuth reagents may be found in Barton,D.H.E., et al., J. Chem. Soc. Chem. Commun., 1986, 65 and referencescited therein.

Similarly, the 31-O-aryl compounds may be prepared by protecting the42-alcohol of rapamycin with a protecting group, such as with atert-butyl dimethylsilyl group, followed by arylation of the 31-positionwith a triaryl bismuth reagent. Removal of the protecting group providesthe 31-O-aryl compounds. In the case of the tert-butyl dimethylsilylprotecting group, deprotection can be accomplished under mildly acidicconditions.

If desired, the 31-hydroxy-42-O-aryl rapamycin, or 31-O-aryl-42-hydroxyrapamycin may be treated with a different triarylbismuth diacetatereagent (prepared immediately prior to use by procedures analogous tothose disclosed above), to give mixed 31-O-aryl-42-O-aryl macrolides.

Reaction Scheme C

As shown in Reaction Scheme C, a solution of the rapamycin in an inertorganic solvent such as methylene chloride, chloroform, pentane, hexane,cyclohexane, heptane or the like or mixtures thereof is treated with analkyl, alkenyl or alkynyl trichloroacetimidate reagent (prepared by thereaction of an appropriate sodium alkoxide with trichloroacetonitrile,such as described by Wessel, H.P., Iversen, T., Bundle, D.R., J. Chem.Soc., Perkin Trans. I, 1985, 2247) in the presence of a mild acidcatalyst such as trifluoromethanesulfonic acid, p-toluenesulfonic acid,methane-sulfonic acid, benzenesulfonic acid, p-nitrobenzene-sulfonicacid, p-bromobenzenesulfonic acid, p-chlorobenzenesulfonic acid, orp-methoxybenzenesulfonic acid, or mixtures thereof at a temperature of20°-50° C.,preferably room temperature, for a period of one hour toseven days, preferably one day, to give the 31- and/or 42-O-alkyl,-alkenyl or -alkynyl rapamycin derivative.

In addition, the procedure of Reaction Schemes A, B, and C may becombined to produce rapamycin derivatives bearing O-aryl, O-alkyl,O-alkenyl and/or O-alkynyl substituents at the 31 and 42 positions.

The procedures described in Reaction Scheme B may be conducted on themonosubstituted products of Reaction Scheme C (and visa versa) to obtainthe mixed disubstituted compounds. In fact, within Reaction Schemes Band C, treatment of the monosubstituted product with a different reagentwill afford the mixed disubstituted compounds.

Reaction Scheme E

As shown in Reaction Scheme E, the 42-hydroxy-3l-R² O -macrolide oralternatively the 31-hydroxy-42-R¹ O-macrolide (not depicted) (whereinR³ is protected hydroxy or hydrogen) may be reacted with an alkenyltrichloroacetimidate (wherein R¹ is C₃₋₁₀ alkenyl) under conditionsdescribed in Reaction Scheme C to give the C-42-O-alkenyl macrolide.Treatment with a stoichiometric amount of osmium tetraoxide in an inertorganic solvent, such as diethyl ether or tetrahydrofuran, in thepresence of an amine base, such as pyridine or 4-methylmorpholineN-oxide, at or near room temperature gives the corresponding glycol.Treatment of the glycol with sodium metaperiodate in a solution oftetrahydrofuran/water gives the aldehyde (wherein A is C₁₋₈ alkyl).Alternatively, the alkenyl macrolide may be treated with sodiummetaperiodate in the presence of a catalytic amount of osmium tetroxidein an organic solvent to give the aldehyde directly. The aldehyde may befurther oxidized to the carboxylic acid by treatment with sodiumchlorite in buffered, aqueous tert-butanol.

Reaction Scheme F

A variety of compounds may be prepared from the corresponding aldehydeas illustrated in Reaction Scheme F. The aldehyde may be reacted with aprimary or secondary amine (wherein R⁶ and R⁷ are as defined above) inan organic solvent such as tetrahydrofuran to give an imine which isreduced in situ with a hydride reducing agent, such as potassiumtriphenyl borohydride or sodium cyanoborohydride, to give the macrolidebearing an amino alkoxy functionality at C-42. The aldehyde may also bereduced to the corresponding alcohol by treatment with a hydridereducing agent, such as potassium triphenyl borohydride or sodiumcyanoborohydride in an organic solvent such as tetrahydrofuran. Thealcohol may be further modified by utilizing the methods of ReactionScheme B (wherein R¹ is unsubstituted or substituted phenyl, naphthyl orbiphenyl) or Reaction Scheme F (wherein R¹ is unsubstituted orsubstituted alkyl, alkenyl or alkynyl) to give the corresponding ether.The procedures described in Reaction Scheme F are readily applicable tothe preparation of compounds bearing analogous functionality at C-31.

Reaction Scheme G

Amide derivatives may be prepared from the carboxylic acid asillustrated in Reaction Scheme G. The carboxylic acid may be coupledwith a primary or secondary amine, HNR⁶ R⁷ (wherein R⁶ and/or R⁷ are asdefined) by any of the peptide coupling methods commonly used in theart, such as with BOP reagent, DCC/HOBT, or EDC/HOBT.

Reaction Scheme H

Hydroxy and keto derivatives may be prepared from the correspondingaldehyde as illustrated in Reaction Scheme H. The aldehyde is reactedwith a nucleophilic organometallic reagent such as a Grignard reagent,an organolithium reagent, or an organocerium reagent in an organicsolvent such as methylene chloride or tetrahydrofuran to give thesubstituted hydroxy compound. Removal of hydroxy protecting groups atother positions of the macrolide (if necessary) gives the macrolidebearing a substituted hydroxy alkoxy functionality at C-42. The alcoholmay also be oxidized to the corresponding ketone by well known methods,such as with 4-methylmorpholine-N-oxide in the presence oftetrapropylammonium perruthenate catalyst under dehydrative conditions.Removal of hydroxy protecting groups (if necessary) gives the macrolidebearing a substituted keto alkoxy functionality at C-42. The proceduresdescribed in Reaction Scheme H are readily applicable to the preparationof compounds bearing analogous functionality at C-31.

Reaction Scheme I

Hydroxy macrolides (wherein R¹ and/or R² bear a hydroxy group) may befurther derivatized by alkylation, acylation or phosphorylation to giveether, ester or phosphate derivatives (wherein R¹ and/or R² bear an--OR¹¹ as defined above) by procedures well known to the practitioner ofthe art.

The object compounds of Formula I obtained according to the reactions asexplained above can be isolated and purified in a conventional manner,for example, extraction, precipitation, fractional crystallization,recrystallization, chromatography, and the like.

It is to be noted that in the aforementioned reactions and thepost-treatment of the reaction mixture therein, the stereoisomer(s) ofstarting and object compounds due to asymmetric carbon atom(s) or doublebond(s) of the object compounds of Formula I may occasionally betransformed into the other stereoisomer(s), and such cases are alsoincluded within the scope of the present invention.

In the present invention, compounds with asymmetric centers may occur asracemates, as diastereomeric mixtures and as individual diastereomers,with all isomeric forms of the compounds being included in the presentinvention.

The compounds of the present invention are capable of forming salts withvarious inorganic and organic acids and bases and such salts are alsowithin the scope of this invention. Examples of such acid addition salts(which are negative counterions defined herein as M⁻) include acetate,adipate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate,camphorate, camphorsulfonate, ethanesulfonate, fumarate, hemisulfate,heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide,methanesulfonate, lactate, maleate, methanesulfonate,2-naphthalenesulfonate, oxalate, pamoate, persulfate, picrate, pivalate,propionate, succinate, tartrate, tosylate, and undecanoate. Base salts(which are positive counterions defined herein as M⁺) include ammoniumsalts, alkali metal salts such as sodium, lithium and potassium salts,alkaline earth metal salts such as calcium and magnesium salts, saltswith organic bases such as dicyclohexylamine salts,N-methyl-D-glucamine, and salts with amino acids such as arginine,lysine and so forth. Also, the basic nitrogen-containing groups may bequaternized with such agents as: lower alkyl halides, such as methyl,ethyl, propyl, and butyl chloride, bromides and iodides; dialkylsulfates like dimethyl, diethyl, dibutyl; diamyl sulfates; long chainhalides such as decyl, lauryl, myristyl and stearyl chlorides, bromidesand iodides; aralkyl halides like benzyl bromide and others. Thenon-toxic physiologically acceptable salts are preferred, although othersalts are also useful, such as in isolating or purifying the product.

The salts may be formed by conventional means, such as by reacting thefree base form of the product with one or more equivalents of theappropriate acid in a solvent or medium in which the salt is insoluble,or in a solvent such as water which is removed in vacuo or by freezedrying or by exchanging the anions of an existing salt for another anionon a suitable ion exchange resin.

C. Utility of the compounds within the scope of the invention

The compounds of Formula I may be employed as immunosuppressants orantimicrobial compounds by methods and in dosages known in the prior artfor rapamycin. These compounds possess pharmacological activity such asimmunosuppressive activity, antimicrobial activity, and the like, andtherefore are useful for the treatment and prevention of the resistanceto transplantation or transplantation rejection of organs or tissuessuch as heart, kidney, liver, duodenum, small-bowel, medulla ossium,skin, pancreatic islet-cell, etc., graft-versus-host diseases by medullaossium transplantation, autoimmune diseases such as rheumatoidarthritis, systemic lupus erythematosis, Hashimoto's thyroiditis,multiple sclerosis, myasthenia gravis, type I diabetes, uveitis,allergic encephalomyelitis, glomerulonephritis, etc., and infectiousdiseases caused by pathogenic microorganisms, particularly fungalinfections.

The compounds of Formula I are also useful for treating or preventinginflammatory and hyperproliferative skin diseases and cutaneousmanifestations of immunologically-mediated illnesses such as: psoriasis,atopical dermatitiis, contact dermatitis and further eczematousdermatitises, seborrhoeic dermatitis, Lichen planus, Pemphigus, bullousPemphigoid, Epidermolysis bullosa, urticaria, angioedemas, vasculitides,erythemas, acne, cutaneous eosinophilias or Alopecia areata. Moreparticularly, the compounds of Formula I are useful in hairrevitalizing, such as in the treatment or prevention of male patternalopecia or alopecia senilis, by providing epilation prevention, hairgermination, and/or a promotion of hair generation and hair growth.

The compounds of Formula I are further useful for treating or preventingreversible obstructive airways disease, including conditions such asasthma, including bronchial asthma, allergic asthma, intrinsic asthma,extrinsic asthma and dust asthma, particularly chronic or inveterateasthma (for example late asthma and airway hyperresponsiveness),bronchitis and the like. The compounds of Formula I may also be usefulfor treating hepatic injury associated with ischemia.

The compounds of Formula I are also useful for treating multidrugresistance of tumor cells, (i.e. enhancing the activity and/orsensitivity of chemotherapeutic agents), preventing or treatinginflammation of mucosa or blood vessels, LTB₄ -mediated diseases,gastric ulcers, vascular damage caused by ischemic diseases andthrombosis, ischemic bowel disease, inflammatory bowel disease (e.g.,Crohn's disease and ulcerative colitis) necrotizing enterocolitis, orintestinal lesions associated with thermal burns, cytomegalovirusinfection, particularly HCMV infection, idiopathic thrombocytopenicpurpura and Basedow's disease.

Further, the compounds of Formula I are also useful for treating orpreventing renal diseases selected from interstitial nephritis,Goodpasture's syndrome, hemolytic-uremic syndrome and diabeticnephropathy; nervous diseases selected from multiple myositis,Guillain-Barre syndrome, Meniere's disease and radiculopathy; endocrinediseases selected from hyperthyroidism; hematic diseases selected frompure red cell aplasia, aplastic anemia, hypoplastic anemia, autoimmunehemolytic anemia, agranulocytosis and anerythroplasia; bone diseasessuch as osteoporosis; respiratory diseases selected from sarcoidosis,fibroid lung and idiopathic interstitial pneumonia; eye diseasesselected from herpetic keratitis, conical cornea, dystrophiaepithelialis corneas, corneal leukmas, ocular pemphigus, Mooren's ulcer,scleritis and Gravels ophthalmopathy skin diseases selected fromdermatomyositis, leukoderma vulgaris, ichthyosis vulgaris, photoallergicsensitivity and cutaneous T cell lymphoma; circulatory diseases selectedfrom arteriosclerosis, aortitis syndrome, polyarteritis nodosa andmyocardosis; collagen diseases selected from scleroderma, Wegener'sgranuloma and Sjogren's syndrome; adiposis; eosinophilic fasciitis;periodontal disease; and muscular dystrophy.

The compounds of Formula I are useful for the treatment of fungalinfections in animals, especially mammals, including humans, inparticular humans and domesticated animals (including farm animals). Thecompounds may be used for the treatment of topical fungal infections inman caused by, among other organisms, species of Candida, Trichophyton,Microsporum or Epidermophyton or in mucosal infections caused by CandidaAlbicans (e.g. thrush and vaginal candidiasis). They may also be used inthe treatment of systemic fungal infections caused by, for exampleCandida albicans, Cryptococcus neoformans, Aspergillus fumigatus,Coccidiodes, Paracocciciodes, Histoplasma or Blastomyces spp. They mayalso be of use in treating eumycotic mycetoma, chromoblastomycosis andphycomycosis.

The pharmaceutical compositions of this invention can be used in theform of a pharmaceutical preparation, for example, in solid, semisolidor liquid form, which contains one or more of the compounds of thepresent invention, as an active ingredient, in admixture with an organicor inorganic carrier or excipient suitable for external, enteral orparenteral applications. The active ingredient may be compounded, forexample, with the usual nontoxic, pharmaceutically acceptable carriersfor tablets, pellets, capsules, suppositories, solutions, emulsions,suspensions, and any other form suitable for use. The carriers which canbe used are water, glucose, lactose, gum acacia, gelatin, mannitol,starch paste, magnesium trisilicate, talc, corn starch, keratin,colloidal silica, potato starch, urea and other carriers suitable foruse in manufacturing preparations, in solid, semisolid, or liquid form,and in addition auxiliary, stabilizing, thickening and coloring agentsand perfumes may be used. The active object compound is included in thepharmaceutical composition in an amount sufficient to produce thedesired effect upon the process or condition of diseases.

For the treatment of these conditions and diseases caused byimmmunoirregularity a compound of formula I may be administered orally,topically, parenterally, by inhalation spray or rectally in dosage unitformulations containing conventional non-toxic pharmaceuticallyacceptable carriers, adjuvants and vehicles. The term parenteral as usedherein includes subcutaneous injections, intravenous, intramuscular,intrasternal injection or infusion techniques.

Dosage levels of the compounds of the present invention are of the orderfrom about 0.005 mg to about 50 mg per kilogram of body weight per day,preferably from about 0.1 mg to about 10 mg per kilogram of body weightper day, are useful in the treatment of the above-indicated conditions(from about 0.7 mg to about 3.5 g per patient per day, assuming a 70 kgpatient). In addition, the compounds of the present invention may beadministered on an intermittent basis; i.e. at daily, semiweekly,weekly, semi-monthly or monthly intervals.

The amount of active ingredient that may be combined with the carriermaterials to produce a single dosage form will vary depending upon thehost treated and the particular mode of administration. For example, aformulation intended for the oral administration of humans may containfrom 0.5 mg to 5 gm of active agent compounded with an appropriate andconvenient amount of carrier material which may vary from about 5 toabout 95 percent of the total composition. Dosage unit forms willgenerally contain from about 0.5 mg to about 500 mg of activeingredient, and preferably about 0.5 mg to about 100 mg of activeingredient. For external administration the compound of Formula I may beformulated within the range of, for example, 0.0001% to 60% by weight,preferably from 0.001 to 10% by weight, and most preferably from about0.005 to 0.8% by weight.

It will be understood, however, that the specific dose level for anyparticular patient will depend on a variety of factors including theactivity of the specific compound employed, the age, body weight,general health, sex, diet, time of administration, route ofadministration, rate of excretion, drug combination and the severity ofthe particular disease undergoing therapy.

The following examples are given for the purpose of illustrating thepresent invention and shall not be construed as being limitations on thescope or spirit of the instant invention.

EXAMPLE 1 General Procedure for the Preparation of Triarylbismuthines

To a stirred suspension of magnesium (486 mg, 20 mmol) in drytetrahydrofuran (10 mi) is added slowly a solution of aryl halide (20mmol) in dry tetrahydrofuran (10 mi). If necessary the mixture is warmedgently to effect Grignard formation. To the stirred solution of theGrignard reagent is added a solution of bismuth trichloride (1.9 g, 6mmol) dissolved in dry tetrahydrofuran (20 mi). The resulting mixture isstirred for 24 hours. The reaction mixture is poured into a separatoryfunnel containing brine and extracted 4× with CH₂ Cl₂. The organicextracts were combined-and dried over anhydrous Na₂ SO₄. The mixture wasfiltered and concentrated in vacuo. The triarylbismuthine is isolatedand purified by flash column chromatography on silica gel.

EXAMPLE 2 43-O-phenyl-rapamycin

To a stirred solution of triphenylbismuthine (100 mg, 0.11 mmol) in CH₂Cl₂ (4 mL) was added peracetic acid (0.041 mL, 0.19 mmol, 32 wt% indilute acetic acid). To this stirred solution was added THF (1 mL),rapamycin (100 mg, 0.126 mmol) and Cu(OAc)₂ (6 mg, 0.03 mmol) and thereaction mixture was stirred at room temperature for 1 hr. The flask wasfitted with a reflux condenser and the mixture was heated to 40° C. for2 hours. The mixture was allowed to cool and was quenched with saturatedaqueous NaHCO₃ and extracted 4× with CH₂ Cl₂. The organic extracts werecombined and dried over anhydrous Na₂ SO₄. The mixture was filtered andconcentrated in vacuo. The products were isolated by preparative TLC onsilica gel (eluted with 2:1 hexanes/acetone) to afford 54.7 mg42-O-phenyl-rapamycin. (¹ H NMR and mass spectral analysis wereconsistent with the desired structure).

EXAMPLE 3 T-Cell Proliferation Assay

1. Sample Preparation

The compounds to be assayed were dissolved in absolute ethanol at 1mg/ml.

2. Assay

Spleens from C57Bl/6 mice were taken under sterile conditions and gentlydissociated in ice-cold RPMI 1640 culture medium (GIBC), Grand Island,N.Y.) supplemented with 10% heat-inactivated fetal calf serum (GIBO)).Cells were pelleted by centrifugation at 1500 rpm for 8 minutes.Contaminating red cells were removed by treating the pellet withammonium chloride lysing buffer (GIBO)) for 2 minutes at 4° C. Coldmedium was added and cells were again centrifuged at 1500 rpm for 8minutes. T lymphocytes were then isolated by separation of the cellsuspension on nylon wool columns as follows: Nylon wool columns wereprepared by packing approximately 4 grams of washed and dried nylon woolinto 20 ml plastic syringes. The columns were sterilized by autoclavingat 25° F. for 30 minutes, Nylon wool columns were wetted with warm (37°C.) culture medium and rinsed with the same medium. Washed spleen cellsresuspended in warm medium were slowly applied to the nylon wool. Thecolumns were then incubated in an upright position at 37° C. for 1 hour.Non-adherent T lymphocytes were eluted from the columns with warmculture medium and the cell suspensions were spun as above.

Purified T lymphocytes were resuspended at 2.5×10⁵ cells/ml in completeculture medium composed of RPMI 1640 medium with 10% heat-inactivatedfetal calf serum, 100 mM glutamine, 1 MM sodium pyruvate, 2×10⁻⁵ M2-mercaptoethanol and 50 μg/ml gentamycin. Ionomycin was added at 250ng/ml and PMA at 10 ng/ml. The cell suspension was immediatelydistributed into 96 well flat-bottom microculture plates (Costar) at 200μl/well. The various dilutions of the compound to be tested were thenadded in triplicate wells at 20 μl/well. The compound17-allyl-1,14-dihydroxy-12-[2'-(4"-hydroxy-3"-methoxycyclohexyl)-1'-methylvinyl]-23,25-dimethoxy-13,19,21,27-tetramethyl-11,28-dioxa-4-aza-tricyclo[22.3.1.0⁴,9octacos-18-ene-2,3,10,16-tetraone was used as a standard. The cultureplates were then incubated at 37° C. in a humidified atmosphere of 5%CO₂ -95% air for 44 hours. The proliferation of T lymphocytes wasassessed by measurement of tritiated thymidine incorporation. After 44hours of culturing, the cells were pulse-labelled with 2 μci/well oftritiated thymidine (NEN, Cambridge, Mass.). After another 4 hours ofincubation, cultures were harvested on glass fiber filters using amultiple sample harvester. Radioactivity of filter discs correspondingto individual wells was measured by standard liquid scintillationcounting methods (Betacounter). Mean counts per minute of replicatewells were calculated and the results expressed as concentration ofcompound required to inhibit tritiated thymidine uptake of T-cells by50%.

A selection of compounds were tested according to the previousprocedure. The title compound of the following Example had activity ininhibiting the proliferation of T-cells in the aforementioned assay: 2.

While the foregoing specification teaches the principles of the presentinvention, with examples provided for the purpose of illustration, itwill be understood that the practice of the invention encompasses all ofthe casual variations, adaptations, modifications, deletions, oradditions of procedures and protocols described herein, as come withinthe scope of the following claims and its equivalents.

What is claimed is:
 1. A compound of formula I: ##STR12## or apharmaceutically acceptable salt thereof, wherein: R¹ and R² areindependently selected from:(1) hydrogen; (2) phenyl; (3) substitutedphenyl in which the substituents are X, Y and Z; (4) 1- or 2- naphthyl;(5) substituted 1- or 2- naphthyl in which the substituents are X, Y andZ; (6) biphenyl; (7) substituted biphenyl in which the substituents areX, Y and Z; (8) C₁₋₁₀ alkyl; (9) substituted C₁₋₁₀ alkyl in which one ormore substituent(s) is(are) selected from:(a) hydroxy, (b) oxo, (c) C₁₋₆-alkoxy, (d) phenyl-C₁₋₃ alkoxy, (e) substituted phenyl-C₁₋₃ alkoxy, inwhich the substituents on phenyl are X, Y and Z, (f) --OCO--CL alkyl,(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are independently selected from(i)hydrogen, (ii) C₁₋₁₀ alkyl unsubstituted or substituted with one or moreof the substituent(s) selected from:(a') phenyl, which is unsubstitutedor substituted with X, Y and Z, (b') --OH, (c') C₁₋₆ alkoxy, (d') --CO₂H) (e') --CO₂ -C₁₋₆ alkyl, (f') --C₃₋₇ cycloalkyl, and (g') --OR¹¹,(iii)C₃₋₁₀ alkenyl unsubstituted or substituted with one or more of thesubstituent(s) selected from:(a') phenyl, which is unsubstituted orsubstituted with X, Y and Z, (b') --OH, (C') C₁₋₆ alkoxy, (d') --CO₂ H,(e') --CO₂ --C₁₋₆ alkyl, (f') --C₃₋₇ cycloalkyl, and (g') --OR¹¹, (iv)orwhere R⁶ and R⁷ and the N to which they are attached can form a3-7-membered saturated heterocyclic ring, unsubstituted or substitutedwith C₁₋₆ alkyl or phenyl, the ring being selected from the groupconsisting of: aziridine, morpholine, thiomorpholine,thiomorpholine-oxide, thiomorpholine-dioxide, piperidine, pyrrolidine,and piperizine, (h) --NR⁶ CO--C₁₋₆ alkyl--R⁷, wherein R⁶ is as definedabove, (i) --NR₆ CO₂ --C₁₋₆ alkyl--R⁷, (j) --NR⁶ CONR⁶ R⁷, (k) --OCONR₆R₇, (l) --COOR₆, (m) --CHO, (n) phenyl, (o) substituted phenyl in whichthe substituents are X, Y and Z, (p) phenyloxy, (q) substitutedphenyloxy in which the substituents are X, Y and Z, (r) 1- or 2-naphthyl, (s) substituted 1- or 2- naphthyl in which the substituentsare X, Y and Z, (t) biphenyl (u) substituted biphenyl in which thesubstituents are X, Y and Z; (v) --OR¹¹, and (w) --S(O)_(p) --C₁₋₆alkyl; (10) C₃₋₁₀ alkenyl; (11) substituted C₃₋₁₀ alkenyl in which oneor more substituent(s) is(are) selected from:(a) hydroxy, (b) oxo, (c)C₁₋₆ alkoxy, (d) phenyl-C₁₋₃ alkoxy, (e) substituted phenyl-C₁₋₃ alkoxy,in which the substituents on phenyl are X, Y and Z, (f) --OCO--C₁₋₆alkyl, (g) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above (h) --NR⁶CO--C₁₋₆ alkyl, wherein R⁶ is as defined above, (i) --COOR⁶, wherein R⁶is as defined above, (j) --CHO, (k) phenyl, (l) substituted phenyl inwhich the substituents are X, Y and Z, (m) 1- or 2-naphthyl, (n)substituted 1- or 2-naphthyl in which the substituents are X, Y and Z,(o) biphenyl, (p) substituted biphenyl in which the substituents are X,Y and Z, (q) --OR¹¹, and (r) --S(O) p--C₁₋₆ alkyl; (12) C₃₋₁₀ alkynyl;(13) substituted C₃₋₁₀ alkynyl in which one or more substituent(s)is(are) selected from:(a) hydroxy, (b) oxo, (c) C₁₋₆ alkoxy, (d)phenyl-C alkoxy, (e) substituted phenyl-C₁₋₃ alkoxy, in which thesubstituents on phenyl are X, Y and Z, (f) --OCO--C₁₋₆ alkyl, (g) --NR₆R , wherein R and R are as defined above, (h) --NR₆ CO--C₁₋₆ alkyl,wherein R⁶ is as defined above, (i) --COOR⁶, wherein R⁶ is as definedabove, (j) --CHO, (k) phenyl, (l) substituted phenyl in which thesubstituents are X, Y and Z, (m) 1- or 2-naphthyl, (n) substituted 1- or2-naphthyl in which the substituents are X, Y and Z, (o) biphenyl, (p)substituted biphenyl in which the substituents are X, Y and Z, and (q)--OR¹¹ ; with the proviso that R¹ and R² are not simultaneouslyhydrogen; R¹¹ is selected from:(a) --PO(OH)O⁻ M⁺, wherein M⁺ is apositively charged inorganic or organic counterion, (b) --SO₃ ⁻ M⁺, (c)--CO(CH₂)_(q) CO₂ ⁻ M⁺, wherein q is 1-3, and (d) --CO--C₁₋₆ alkyl-NR⁶R⁷, wherein R⁶ and R⁷ are as defined above and the alkyl isunsubstituted or substituted with one or more substituents selectedfrom:(i) hydroxy, (ii) C₁₋₆ alkoxy, (iii) --NR¹⁶ R¹⁷, wherein R¹⁶ andR¹⁷ are independently selected from:(a') hydrogen, and (b') C₁₋₆ alkyl,(iv) --COOR⁶, wherein R⁶ is as defined above, (v) phenyl, (vi)substituted phenyl in which the substituents are X, Y and Z, (vii) --SH,and (viii) --S--C₁₋₆ alkyl; X, Y and Z independently are selectedfrom:(a) hydrogen, (b) C₁₋₇ alkyl, (c) C₂₋₆ alkenyl, (d) halogen, (e)--(CH₂)_(m) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above, and m is 0to 2, (f) --CN, (g) --CHO, (h) --CF₃) (i) --SR⁸, wherein R⁸ is hydrogen,C₁₋₆ alkyl, trifluoromethyl, or phenyl, (j) --SOR⁸, wherein R⁸ is asdefined above, (k) --SO₂ R⁸, wherein R⁸ is as defined above, (l) --CONR⁶R⁷, wherein R⁶ and R⁷ are as defined above, (m) R⁹ O(CH₂)_(m) - whereinR⁹ is hydrogen, C₁₋₃ alkyl, hydroxy-C₂₋₃ alkyl, trifluoromethyl, phenylor naphthyl and m is as defined above, (n) --CH(OR¹²)(OR¹³), wherein R¹²and R¹³ are C₁₋₃ alkyl or taken together form an ethyl or propyl bridge,(o) ##STR13## wherein R⁹ and m are as defined above, and (p) ##STR14##wherein R⁹ and m are as defined above, and (q) --OR¹¹ ; or any two ofadjacent X, Y and Z can be joined to form a ring selected from the groupconsisting of: dioxolanyl, dihydrofuranyl, dihydropyranyl, and dioxanyl.2. The compound of claim 1 wherein:R¹ and R² are independently selectedfrom:(1) hydrogen; (2) methyl; (3) phenyl; (4) substituted phenyl inwhich the substituents are X, Y and Z; (5) 1- or 2- naphthyl; (6)substituted 1- or 2- naphthyl in which the substituents are X, Y and Z;(7) biphenyl; and (8) substituted and biphenyl in which the substituentsare X, Y and Z; with the proviso that R¹ and R² are not simultaneouslyhydrogen; X, Y and Z are independently, selected from:(a) hydrogen, (b)C₁₋ 7 alkyl, (c) C₂₋₆ alkenyl, (d) halogen, (e) --(CH₂)_(m) -NR⁶ R⁷,wherein R⁶ and R⁷ are, independently selected from(i) hydrogen, or (ii)C₁₋₆ alkyl unsubstituted or substituted with phenyl, and m is 0 to 2,(f) --CN, (g) --CHO, (h) --CF₃, (i) --SR⁸, wherein R⁸ is hydrogen, C₁₋₆alkyl, trifluoromethyl, or phenyl, (j) --SOR⁸, wherein R⁸ is as definedabove, (k) --SO₂ R⁸, wherein R⁸ is as defined above, (l) --CONR⁶ R⁷,wherein R⁶ and R⁷ are as defined above, (m) R⁹ O(CH₂)_(m) - wherein R⁹is hydrogen, C₁₋₃ alkyl, hydroxy-C₂₋₃ alkyl, trifluoromethyl, phenyl ornaphthyl and m is as defined above, (n) --CH(OR¹²)(OR¹³), wherein R¹²and R¹³ are C₁₋₃ alkyl or taken together form an ethyl or propyl bridge,(o) ##STR15## wherein R⁹ and m are as defined above, and (p) ##STR16##wherein R⁹ and m are as defined above, and (q) --OR¹¹ ; or any two ofadjacent X, Y and Z can be joined to form a ring selected from the groupconsisting of: dioxolanyl, dihydrofuranyl, dihydropyranyl, and dioxanyl.3. The compound of claim 1 wherein:R¹ and R² are independently selectedfrom:(1) hydrogen; (2) C₁₋₁₀ alkyl; (3) substituted C₁₋₁₀ alkyl in whichone or more substituent(s) is(are) selected from:(a) hydroxy, (b) oxo,(c) C₁₋₆ -alkoxy, (d) phenyl-C₁₋₃ alkoxy, (e) substituted phenyl-C₁₋₃alkoxy, in which the substituents on phenyl are X, Y and Z, (f)--OCO--C₁₋₆ alkyl, (g) --NR⁶ R⁷, wherein R⁶ and R⁷ are independentlyselected from(i) hydrogen, (ii) C₁₋₁₀ alkyl unsubstituted or substitutedwith one or more of the substituent(s) selected from:(a') phenyl, whichis unsubstituted or substituted with X, Y and Z, (b') --OH, (c') C₁₋₆alkoxy, (d') --CO₂ H, (e') --CO₂ --C₁₋₆ alkyl, (f') --C₃₋₇ cycloalkyl,and (g') --OR¹¹, (iii)C₃₋₁₀ alkenyl unsubstituted or substituted withone or more of the substituent(s) selected from:(a') phenyl, which isunsubstituted or substituted with X, Y and Z, (b') --OH, alkoxy, (c')C₁₋₆ alkoxy, (d') --CO₂ H, (e') --CO₂ --C₁₋₆ alkyl, (f') --C₃₋₇cycloalkyl, and (g') --OR¹¹, (iv)or where R⁶ and R⁷ and the N to whichthey are attached can form a 3-7-membered saturated heterocyclic ring,unsubstituted or substituted with C₁₋₆ alkyl or phenyl, the ring beingselected from the group consisting of: aziridine, morpholine,thiomorpholine, thiomorpholine-oxide, thiomorpholine-dioxide,piperidine, pyrrolidine, and piperazine, (h) --NR⁶ CO--C₁₋₆ alkyl-R⁷,wherein R⁶ is as defined above, (i) --NR⁶ CO₂ --C₁₋₆ alkyl-R⁷, (j) --NR⁶CONR⁶ R⁷, (k) --OCONR⁶ R⁷, (l) --COOR⁶, (m) --CHO, (n) phenyl, (o)substituted phenyl in which the substituents are X, Y and Z, (p)phenyloxy, (q) substituted phenyloxy in which the substituents are X, Yand Z, (r) 1- or 2- naphthyl, (s) substituted 1- or 2- naphthyl in whichthe substituents are X, Y and Z, (t) biphenyl (u) substituted biphenylin which the substituents are X, Y and Z; (v) --OR¹¹, and (w) --S(O)_(p)--C₁₋₆ alkyl; (4) C₃₋₁₀ alkenyl; (5) substituted C₃₋₁₀ alkenyl in whichone or more substituent(s) is(are) selected from:(a) hydroxy, (b) oxo,(c) C₁₋₆ alkoxy, (d) phenyl-C₁₋₃ alkoxy, (e) substituted phenyl-C₁₋₃alkoxy, in which the substituents on phenyl are X, Y and Z, (f)--OCO--C₁₋₆ alkyl, (g) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above(h) --NR⁶ CO--C₁₋₆ alkyl, wherein R⁶ is as defined above, (i) --COOR⁶,wherein R⁶ is as defined above, (j) --CHO, (k) phenyl, (l) substitutedphenyl in which the substituents are X, Y and Z, (m) 1- or 2-naphthyl,(n) substituted 1- or 2-naphthyl in which the substituents are X, Y andZ, (o) biphenyl, (p) substituted biphenyl in which the substituents areX, Y and Z, (q) --OR¹¹, and (r) --S(O)_(p) --C₁₋₆ alkyl; (6) C₃₋₁₀alkynyl; (7) substituted C₃₋₁₀ alkynyl in which one or moresubstituent(s) is(are) selected from:(a) hydroxy, (b) oxo, (c) C₁₋₆alkoxy, (d) phenyl-C₁₋₃ alkoxy, (e) substituted phenyl-C₁₋₃ alkoxy, inwhich the substituents on phenyl are X, Y and Z, (f) --OCO--C₁₋₆ alkyl,(g) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above, (h) --NR⁶ CO--C₁₋₆alkyl, wherein R⁶ is as defined above, (i) --COOR⁶, wherein R⁶ is asdefined above, (j) --CHO, (k) phenyl, (l) substituted phenyl in whichthe substituents are X, Y and Z, (m) 1- or 2-naphthyl, (n) substituted1- or 2-naphthyl in which the substituents are X, Y and Z, (o) biphenyl,(p) substituted biphenyl in which the substituents are X, Y and Z, and(q) --OR¹¹ ; with the proviso that R¹ and R² are not simultaneouslyhydrogen; R¹¹ is selected from:(a) --PO(OH)O⁻ M⁺, wherein M⁺ is apositively charged inorganic or organic counterion, (b) --SO₃ ⁻ M⁺, (c)--CO(CH₂)_(q) CO₂ ⁻ M⁺, wherein q is 1-3, and (d) --CO--C₁₋₆ alkyl-NR⁶R⁷, wherein R⁶ and R⁷ are as defined above and the alkyl isunsubstituted or substituted with one or more substituents selectedfrom:(i) hydroxy, (ii) C₁₋₆ alkoxy, (iii) --NR¹⁶ R¹⁷, wherein R¹⁶ andR¹⁷ are independently selected from:(a') hydrogen, and (b') C₁₋₆ alkyl,(iv) --COOR⁶, wherein R⁶ is as defined above, (v) phenyl, (vi)substituted phenyl in which the substituents are X, Y and Z, (vii) --SH,and (viii) --S--C₁₋₆ alkyl; X, Y and Z independently are selected from:(a) hydrogen, (b) C₁₋₇ alkyl, (c) C₂₋₆ alkenyl, (d) halogen, (e)--(CH₂)_(m) --NR⁶ R⁷, wherein R⁶ and R⁷ are as defined above, and m is 0to 2, (f) --CN, (g) --CHO, (h) --CF₃, (i) --SR⁸, wherein R⁸ is hydrogen,C₁₋₆ alkyl, trifluoromethyl, or phenyl, (j) --SOR⁸, wherein R⁸ is asdefined above, (k) --SOR₂ R⁸, wherein R⁸ is as defined above, (l)--CONR⁶ R⁷, wherein R⁶ and R⁷ are as defined above, (m) R⁹ O(CH₂)_(m) -wherein R⁹ is hydrogen, C₁₋₃ alkyl, hydroxy-C₂₋₃ alkyl, trifluoromethyl,phenyl or naphthyl and m is as defined above, (n) --CH(OR¹²)(OR¹³),wherein R¹² are C₁₋₃ alkyl or taken together form an ethyl or propylbridge, (o) ##STR17## wherein R⁹ and m are as defined above, and (p)##STR18## wherein R⁹ and m are as defined above, and (q) --OR¹¹ ; or anytwo of adjacent X, Y and Z can be joined to form a ring selected fromthe group consisting of: dioxolanyl, dihydro-furanyl, dihydropyranyl,and dioxanyl.
 4. A compound which is: ##STR19## wherein R¹ and R² areselected from the following combinations of substituents:

    ______________________________________                                        R.sup.1              R.sup.2                                                  ______________________________________                                        a)                                                                                  ##STR20##          H                                                    b)                                                                                  ##STR21##          H                                                    c)                                                                                  ##STR22##          H                                                    d)                                                                                  ##STR23##          H                                                    e)                                                                                  ##STR24##          H                                                    f)                                                                                  ##STR25##          H                                                    g)                                                                                  ##STR26##          H                                                    h)                                                                                  ##STR27##          H                                                    i)                                                                                  ##STR28##          H                                                    j)   H                                                                                                  ##STR29##                                           k)   H                                                                                                  ##STR30##                                           l)   H                                                                                                  ##STR31##                                           m)                                                                                  ##STR32##          H                                                    n)                                                                                  ##STR33##          H                                                    o)                                                                                  ##STR34##          H                                                    p)                                                                                  ##STR35##          H                                                    q)                                                                                  ##STR36##          H                                                    r)                                                                                  ##STR37##           H.                                                  ______________________________________                                    


5. A compound which is: ##STR38## wherein R¹ and R² are selected fromthe following combinations of substituents:

    ______________________________________                                        R.sup.1           R.sup.2                                                     ______________________________________                                        a)                                                                                   ##STR39##      H                                                       b)    H                                                                                              ##STR40##                                              c)                                                                                   ##STR41##      H                                                       d)    H                                                                                              ##STR42##                                              e)                                                                                   ##STR43##      H                                                       f)    H                                                                                              ##STR44##                                              g)                                                                                   ##STR45##      H                                                       h)                                                                                   ##STR46##      H                                                       i)    H                                                                                              ##STR47##                                              j)                                                                                   ##STR48##      H                                                       k)    H                                                                                              ##STR49##                                              l)                                                                                   ##STR50##      H                                                       m)    H                                                                                              ##STR51##                                              n)                                                                                   ##STR52##      H                                                       o)    H                                                                                              ##STR53##                                              p)                                                                                   ##STR54##      H                                                       q)    H                                                                                              ##STR55##                                              r)                                                                                   ##STR56##      H                                                       s)    H                                                                                              ##STR57##                                              t)                                                                                   ##STR58##      H                                                       u)    H                                                                                              ##STR59##                                              v)                                                                                   ##STR60##      H                                                       w)    H                                                                                              ##STR61##                                              x)                                                                                   ##STR62##      H                                                       y)    H                                                                                              ##STR63##                                              z)                                                                                   ##STR64##      H                                                       aa)                                                                                  ##STR65##      H                                                       bb)                                                                                  ##STR66##      H                                                       cc)                                                                                  ##STR67##      H                                                       dd)                                                                                  ##STR68##      H                                                       ee)                                                                                  ##STR69##      H                                                       ff)                                                                                  ##STR70##      H                                                       gg)                                                                                  ##STR71##      H                                                       hh)   H                                                                                              ##STR72##                                              ii)                                                                                  ##STR73##      H                                                       jj)                                                                                  ##STR74##      H                                                       kk)   H                                                                                              ##STR75##                                              ______________________________________                                    


6. The compound of claim 1 which is: 42-O-phenyl rapamycin.
 7. Apharmaceutical composition comprising a pharmaceutical carrier and atherapeutically effective amount of the compound of claim
 1. 8. A methodfor the treatment of immunoregulatory disorders or diseases comprisingthe administration to a mammalian species in need of such treatment ofan effective amount of the compound of claim
 1. 9. A method for thetreatment of resistance to transplantation comprising the administrationto a mammalian species in need of such treatment of an effective amountof the compound of claim
 1. 10. A method for the treatment ofinflammatory comprising the administration to a mammalian species inneed of such treatment of an effective amount of the compound ofclaim
 1. 11. A method for the treatment of autoimmune diseasescomprising the administration to a mammalian species in need of suchtreatment of an effective amount of the compound of claim
 1. 12. Amethod for the treatment of fungal infections comprising theadministration to a mammalian species in need of such treatment of aneffective amount of the compound of claim 1.